This invention is directed to anti-atherosclerotic agents and more specifically to compounds, compositions and methods for treating atherosclerotic conditions such as dyslipoproteinimias and coronary heart disease. This invention specifically relates to substituted tetrahydro-pyrimidine-2(1H)-thione derivatives that elevate HDL cholesterol (HDL-C) concentration and which may be useful for the treatment of atherosclerotic conditions and coronary heart disease.
Numerous studies have demonstrated that both the risk of coronary heart disease (CHD) in humans and the severity of experimental atherosclerosis in animals are inversely correlated with serum HDL cholesterol (HDL-C) concentrations (Russ et al., Am. J. Med., 11, 480-483 (1951); Gofman et al., Circulation, 34, 679-697 (1966); Miller and Miller, Lancet, 1, 16-19 (1975); Gordon et al., Circulation, 79, 8-15 (1989); Stampfer et al., N. Engl. J. Med. 325, 373-381 (1991); Badimon et al., Lab. Invest., 60, 455-461 (1989)). Atherosclerosis is the process of the accumulation of cholesterol within the arterial wall which results in the occlusion, or stenosis, of coronary and cerebral arterial vessels and subsequent myocardial infarction and stroke. Angiographic studies have shown that elevated levels of some HDL particles in humans appear to be correlated to a decreased number of sites of stenosis in the coronary arteries of humans (Miller et al., Br. Med. J., 282, 1741-1744 (1981)).
There are several mechanisms by which HDL may protect against the progression of atherosclerosis. Studies in vitro have shown that HDL is capable of removing cholesterol from cells (Picardo et al., Arteriosclerosis, 6, 434-441 (1986)). Data of this nature suggest that one antiatherogenic property of HDL may lie in its ability to deplete tissue of excess free cholesterol and eventually lead to the delivery of this cholesterol to the liver (Glomset, J. Lipid Res., 9, 155-167 (1968)). This has been supported by experiments showing efficient transfer of cholesterol from HDL to the liver (Glass et al., J. Biol. Chem., 258 7161-7167 (1983); McKinnon et al., J. Biol. Chem., 26, 2548-2552 (1986)). In addition, HDL may serve as a reservoir in the circulation for apoproteins necessary for the rapid metabolism of triglyceride-rich lipoproteins (Grow and Fried, J. Biol. Chem., 253, 1834-1841 (1978); Lagocki and Scanu, J. Biol. Chem., 255, 3701-3706 (1980); Schaefer et al., J. Lipid Res., 23, 1259-1273 (1982)). Accordingly, agents which increase HDL cholesterol concentrations would be of utility as antiatherosclerotic agents, useful particularly in the treatment of dyslipoproteinimias and coronary heart disease.
Cyclic ureas and thioureas have heretofore been used for various purposes, all of which are unrelated to their antiatherosclerotic effects.
For example, JP 3-176475 discloses the preparation of cyclic ureas and thioureas such as 1,3-disubstituted tetrahydro-pyrimidine-2-thiones and their use as herbicidal agents. European Patent Application Publication Nos. 0612741 and 0503548 disclose cyclic urea (thiourea) derivatives useful as aggregation inhibitors and inhibitors of cell-cell and cell-matrix interactions, respectively.
The J. Chem. Soc. Perkin I, 6, 1622-1625 (1981) describes the regioselective preparation of 3,4-dihydro and 3,4,5,6-tetrahydro-pyrimidine-2(1H)-ones and the corresponding thiones useful as intermediates in the synthesis of diamines, thiazines and pyrimidine-2(1H)-ones. Synthesis, 1175-1180 (1994) describes the synthesis of antiinflammatory pyrimidobenzimidazoles from 1-aryl-6-hydroxy-tetrahydro-pyrimidine-2-(1H)-thiones.
The use of 6-hydroxy-tetrahydro-pyrimidine-2(1H)-thiones as antiamoebic and antihelmintic agents is disclosed in Indian Drugs, 31, 317-320 (1994). Other uses of similar compounds are disclosed in, e.g., Arzneim.-Forsch., 40, 55-57 (1990) (1-phenyl-tetrahydro-pyrimidine-2(1H)-thione as immunomodulator agents); and Farmakol. Toksikol. (Moscow), 41, 494-497 (1978) (1-substituted 4-hydroxy-hexahydro-pyrimidine-2-thione derivatives as radioprotectors). The production of similar thione derivatives, without reference to a specific utility has been disclosed in Dokl. Bolg. Akad. Nauk., 35, 49-51 (1982) (the mass spectra of 1-substituted 2-oxo (or 2-thio)hexahydropyrimidines); Khim. Gerotsikl. Soedin., 1273-1278 (1983) (the mass spectra of 1-substituted 4-hydroxy-hexahydro-pyrimidine-2-thiones; and Khim. Gerotsikl. Soedin., 889-895 (1989) (the synthesis of 4-hydroxy-hexahydro-pyrimidine-2-thiones). None of these references disclose the use of cyclic ureas and thioureas to raise the HDL cholesterol concentrations in mammals.
In accordance with this invention, there are provided 1-(aryl-substituted)-3-substituted tetrahydro-pyrimidine-2(1H)-thiones which are useful as antiatherosclerotic agents.
More particularly, this invention provides antiatherosclerotic agents of Formula 1 having the following structure: 
wherein
R1 is hydrogen, alkyl of 1-6 carbon atoms, cycloalkyl of 3-8 carbon atoms, alkenyl of 2-7 carbon atoms, alkynyl of 2-7 carbon atoms, or phenylalkyl of 7-10 carbon atoms; and
R2, R3, R4, R5, and R6 are each, independently, hydrogen, halogen, alkyl of 1-6 carbon atoms, cycloalkyl of 3-8 carbon atoms, alkenyl of 2-7 carbon atoms, alkynyl of 2-7 carbon atoms, aralkyl of 7-10 carbon atoms, alkoxy of 1-6 carbon atoms, aryloxy of 6-12 carbon atoms, aralkyloxy of 7-12 carbon atoms, fluoroalkoxy of 1-6 carbon atoms, trifluoromethyl, alkylthio of 1-3 carbon atoms, alkylsulfonyl of 1-3 carbon atoms, xe2x80x94SCF3, nitro, alkylamino in which the alkylamino moiety has 1-6 carbon atoms, or dialkylamino in which each alkyl group has 1-6 carbon atoms;
or a pharmaceutically acceptable salt thereof.
This invention also provides methods of elevating the HDL concentration and treating or inhibiting atherosclerosis and related coronary heart disease, or dyslipoproteinemias, and improving the HDL/LDL cholesterol ratio in a mammal in need thereof which comprises administering to the mammal a compound of Formula 1 having the structure: 
wherein
R1 is hydrogen, alky of 1-6 carbon atoms, cycloalkyl of 3-8 carbon atoms, alkenyl of 2-7 carbon atoms, alkynyl of 2-7 carbon atoms, or phenylalkyl of 7-10 carbon atoms; and
R2, R3, R4, R5, and R6 are each, independently, hydrogen, halogen, alkyl of 1-6 carbon atoms, cycloalkyl of 3-8 carbon atoms, alkenyl of 2-7 carbon atoms, alkynyl of 2-7 carbon atoms, aralkyl of 7-10 carbon atoms, alkoxy of 1-6 carbon atoms, aryloxy of 6-12 carbon atoms, aralkyloxy of 7-12 carbon atoms, fluoroalkoxy of 1-6 carbon atoms, trifluoromethyl, alkylthio of 1-3 carbon atoms, alkylsulfonyl of 1-3 carbon atoms, xe2x80x94SCF3, nitro, alkylamino in which the alkylamino moiety has 1-6 carbon atoms, or dialkylamino in which each alkyl group has 1-6 carbon atoms;
or a pharmaceutically acceptable salt thereof.
It is preferred that the compounds of the present invention are represented by the compounds of Formula 1 where R1 is alkyl of 1-6 carbon atoms, alkenyl of 2-7 carbon atoms or cycloalkyl of 3-8 carbon atoms; and R2, R3, R4, R5, and R6 are each, independently, hydrogen, halogen or alkyl of 1-6 carbon atoms.
More preferably, the compounds of the present invention are represented by the compounds of Formula 1 where R1 is methyl, ethyl, isopropyl or cyclobutyl; and R2, R3, R4, R5, and R6 are each, independently, chlorine or methyl.
As used in describing this invention, the terms xe2x80x9calkylxe2x80x9d, xe2x80x9calkenylxe2x80x9d, and xe2x80x9calkynylxe2x80x9d include both straight chain as well as branched moieties. This includes the alkyl portions of substituents such as alkoxy, thioalkyl, alkylsulfinyl, alkylsulfonyl, fluoroalkoxy, and the like. The terms xe2x80x9chaloxe2x80x9d and xe2x80x9chalogenxe2x80x9d include fluorine, chlorine, bromine, and iodine. Fluoroalkoxy includes mono-, di-, tri-, and polyfluorinated alkoxy moieties, such as xe2x80x94OCF3, xe2x80x94OCH2F, xe2x80x94OCHF2, xe2x80x94OCH2CF3, and the like. The term xe2x80x9carylxe2x80x9d includes radicals such as benzyl, phenyl or naphthyl.
As used in describing this invention, the term xe2x80x9ccompounds of this inventionxe2x80x9d includes the broader description encompassing the formula used in accordance with the above methods, as well as the narrower description encompassing the formula used in accordance with the above novel compounds.
The pharmaceutically acceptable salts of the compounds of this invention are those derived from organic and inorganic acids such as, but not limited to: acetic, lactic, citric, tartaric, succinic, fumaric, maleic, malonic, mandelic, malic, hydrochloric, hydrobromic, phosphoric, nitric, sulfuric, methane sulfonic, toluene sulfonic and similarly known acceptable acids.
The most preferred compounds of the present invention are:
1-(4-chloro-2-methyl-phenyl)-3,4,5,6-tetrahydro-pyrimidine-2(1H)-thione;
1-phenyl-3,4,5,6-tetrahydro-pyrimidine-2(1H)-thione;
1-(4-chloro-2-methyl-phenyl)-3-ethyl-3,4,5,6-tetrahydro-pyrimidine-2(1H)-thione;
1-(2,6-dimethyl-phenyl)-3,4,5,6-tetrahydro-pyrimidine-2(1H)-thione;
1-(5-chloro-2-methyl-phenyl)-3-ethyl-3,4,5,6-tetrahydro-pyrimidine-2(1H)-thione;
1-(5-chloro-2-methyl-phenyl)-3-isopropyl-3,4,5,6-tetrahydro-pyrimidine-2(1H)-thione;
1-(4-chloro-2-methyl-phenyl)-3-isopropyl-3,4,5,6-tetrahydro-pyrimidine-2(1H)-thione;
3-allyl-1-(5-chloro-2-methyl-phenyl)-3,4,5,6-tetrahydro-pyrimidine-2(1H)-thione;
1-(5-chloro-2-methyl-phenyl)-3-methyl-3,4,5,6-tetrahydro-pyrimidine-2(1H)-thione;
3-allyl-1-(4-chloro-2-methyl-phenyl)-3,4,5,6-tetrahydro-pyrimidine-2(1H)-thione;
1-(4-chloro-2-methyl-phenyl)-3-methyl-3,4,5,6-tetrahydro-pyrimidine-2(1H)-thione;
3-allyl-1-(6-chloro-2-methyl-phenyl)-3,4,5,6-tetrahydro-pyrimidine-2(1H)-thione;
1-(6-chloro-2-methyl-phenyl)-3-methyl-3,4,5,6-tetrahydro-pyrimidine-2(1H)-thione;
1-(5-chloro-2-methyl-phenyl)-3-isobutyl-3,4,5,6-tetrahydro-pyrimidine-2(1 H)-thione;
1-(6-chloro-2-methyl-phenyl)-3-isopropyl-3,4,5,6-tetrahydro-pyrimidine-2(1H)-thione;
1-(6-chloro-2-methyl-phenyl)-3-ethyl-3,4,5,6-tetrahydro-pyrimidine-2(1H)-thione;
1-(4-chloro-2-methyl-phenyl)-3-isobutyl-3,4,5,6-tetrahydro-pyriniidine-2(1H)-thione;
1-(6-chloro-2-methyl-phenyl)-3-cyclobutyl-3,4,5,6-tetrahydro-pyrimidine-2(1H)-thione;
1-(5-chloro-2-methyl-phenyl)-3-cyclobutyl-3,4,5,6-tetrahydro-pyrimidine-2(1H)-thione; and
1-(6-chloro-2-methyl-phenyl)-3-isobutyl-3,4,5,6-tetrahydro-pyrimidine-2(1H)-thione.
The 1-(aryl-substituted)-3-substituted-tetrahydro-pyrimidine-2(1H)-thiones of this invention may be prepared by cyclocondensation of an appropriately substituted diamine of formula (2) with a thiocarbonylating agent as shown in Scheme 1. 
wherein R1, R2, R3, R4, R5, and R6 are as described above for Formula 1.
The thiocarbonylation agent (3) of Scheme 1 may be thiophosgene in an organic aprotic solvent, such as dichloromethane or chloroform, in the presence of an organic base such as triethylamine, at temperatures ranging from 0xc2x0 C. to ambient essentially according to the method of Sharma et al., J. Med. Chem., 18, 913 (1975) (for a review of thiophosgene in organic syntheses see Sharma, Synthesis, 803 (1978)).
Alternatively, the cyclocondensation can be carried out with a heterocyclic thiocarbonyl transfer reagent (3) such as 1,1xe2x80x2-thiocarbonyldiimidazole (as disclosed in Staab et al., Angew. Chemie, 73, 148 (1961); Staab et al., Justus Liebigs Ann. Chem., 657, 98 (1962); and Larsen et al., J. Org. Chem., 43, 337 (1978)), or 1,1xe2x80x2-thiocarbonyl-di-1,2,4-triazole (as disclosed in Larsen et al., J. Org. Chem., 43, 337 (1978)) in an organic aprotic solvent such as dichloromethane or dioxane at temperatures ranging from ambient to the reflux temperature of the solvent.
It is generally accepted that the cyclocondensation proceeds through the intermediacy of an heterocyclic N-thiocarboxamide such as imidazole- or 1,2,4-triazole-N-thiocarboxamide of general formula (4) as shown in Scheme 2 (Staab, Angew. Chemie Int. Ed., 1, 351 (1962)). It has been found that cyclocondensation of the intermediate (4) to the desired tetrahydro-pyrimidine-2(1H)-thiones of formula (1) can be accelerated by the addition of an organic acid (5) such as para-toluene sulfonic acid monohydrate to the reaction mixture containing the intermediate N-thiocarboxamide of formula (3). 
wherein R1, R2, R3, R4, R5, and R6 are as described above for Formula 1.
The intermediate substituted diamines of general formula (2) of Schemes 1 and 2 can be prepared by either one of the two routes shown in Scheme 3. 
Thus, an appropriately substituted aniline of general formula (6) can be alkylated with a haloalkylamine, preferably a bromo(chloro)alkylamine of formula (7, W=Br, Cl), in the absence of a solvent and at temperatures ranging from ambient to the reflux temperature of the aniline employed, to provide the desired diamines of Formula (2) of Schemes 1 and 2.
Alternatively, the aniline of formula (6) is first allylated with a haloalkanol, preferably a bromo(chloro)alkanol of general formula (8, W=Br, Cl), to provide the intermediate aminoalkanol of formula (9) which is in turn converted to the haloalkylamine intermediate of formula (10, W=Br, Cl). Reaction of (10) with an amine of formula (11) yields the desired diamines of formula (2) of Schemes 1 and 2, wherein R1, R2, R3, R4, R5, and R6 are as described above for formula 1.
The appropriately substituted aniline starting materials (6) of Scheme 3 are either available commercially or can be prepared by procedures analogous to those in the literature for known compounds (see, e.g., J. March, Advanced Organic Chemistry, 3rd Ed., Wiley-Interscience, NY, page 1153).
The preferred bromo(chloro)alkylamines of formula (7) of Scheme 3 can be obtained by halogenation of the corresponding aminoalkanols of formula (12) as shown in Scheme 4 (see, Angiolini et al., Gazz. Chim. Ital., 106, 111 (1976); Crabb et al., Tetrahedron, 26, 3941 (1970); Deady et al., J. Chem. Soc. Perkin I, 782 (1973); Deady et al., J. Org. Chem., 28, 511 (1963); Sammes et al., J. Chem. Soc. Perkin I, 2415 (1984)). 
The preferred aminoalkanols of formula (12) of Scheme 4 are either available commercially or can be prepared by procedures analogous to those in the literature for known compounds (see, e.g., Will et al., Annalen, 568, 34 (1950); Elderfield et al., J. Am. Chem. Soc., 68, 1579 (1946); Angiolini et al., Gazz. Chim. Ital., 106, 111 (1976); Jones et al., J. Chem. Soc. (B), 1300 (1971)).
The preferred bromo(chloro)alkanols of formula (8) of Scheme 3 are either available commercially or are known in the art.
Representative compounds of this invention were evaluated in an in vivo standard pharmacological test procedure which measured the ability of the compounds of this invention to elevate HDL cholesterol levels. The following briefly describes the procedure used and results obtained. Male Sprague-Dawley rats weighing 200-225 g were housed two per cage and fed Purina Rodent Chow Special Mix 5001-S supplemented with 0.25% cholic acid and 1.0% cholesterol and water ad libitum for 8 days. Each test substance was administered to a group of six rats fed the same diet with the test diet mixed in as 0.005-0.1% of the total diet. Body weight and food consumption were recorded prior to diet administration and at termination. Typical doses of the test substances were 5-100 mg/kg/day.
At termination, blood was collected from anesthetized rats and the serum was separated by centrifugation. Total serum cholesterol was assayed using the Sigma Diagnostics enzymatic kit for the determination of cholesterol, Procedure No. 352, modified for use with ninety-six well microtiter plates. After reconstitutiton with water the reagent contains 300 U/l cholesterol oxidase, 100 U/l cholesterol esterase, 1000 U/l horse radish peroxidase, 0.3 mmol/l 4-aminoantipyrine and 30.0 mmol/l p-hydroxybenzene sulfonate in a pH 6.5 buffer. In the reaction cholesterol was oxidized to produce hydrogen peroxide which was used to form a quinoneimine dye. The concentration of dye formed was measured spectrophotometrically by absorbance at 490 nm after incubation at 25xc2x0 C. for 30 minutes. The concentration of cholesterol was determined for each serum sample relative to a commercial standard from Sigma.
HDL cholesterol concentrations in serum were determined by separation of lipoprotein classes by fast protein liquid chromatography (FPLC) by a modification of the method of Kieft et al., (J. Lipid Res., 32, 859-866 (1991). Using this methodology, 25 mL of serum was injected onto Superose 12 and Superose 6 (Pharmacia), in series, with a column buffer of 0.05 M Tris (2-amino-2-hydroxymethyl-1,3-propanediol) and 0.15 M sodium chloride at a flow rate of 0.5 mL/min. The eluted sample was mixed on line with Boehringer-Mannheim cholesterol reagent pumped at 0.2 mL/min. The combined eluents were mixed and incubated on line through a knitted coil (Applied Biosciences) maintained at a temperature of 45xc2x0 C. The eluent was monitored by measuring absorbance at 490 nm and gives a continuous absorbance signal proportional to the cholesterol concentration. The relative concentration for each lipoprotein class was calculated as the percent of total absorbance. HDL cholesterol concentration in serum was calculated as the percent of total cholesterol as determined by FPLC multiplied by the total serum cholesterol concentration.
The results obtained in this standard pharmaceutical test procedure are shown below in Table 1. In Examples 5-7, 9, 10 and 12-20 the test compounds were administered at a dose of 100 mg/kg. In Examples 1, 2, 3 and 11, the test compounds were administered at a dose of 50 mg/kg. In Example 4, the test compound was administered at a dose of 46 mg/kg and in Example 8 at a dose of 90 mg/kg. The duration of treatment for all examples was eight days.
The results set forth in Table I demonstrate that the compounds of this invention are useful in raising the concentration of HDL cholesterol. Therefore, the compounds of this invention are useful for treating or inhibiting atherosclerosis, related cardiovascular disease, or dyslipoproteinemias, and for improving the HDL/LDL cholesterol ratio, and several metabolic conditions associated with low concentrations of HDL such as low HDL-cholesterol levels in the absence of dyslipidemia, metabolic syndrome, non-insulin dependent diabetes mellitus (NIDDM), familial combined hyperlipidemia, familial hypertriglyceridemia, and dyslipidemia in peripheral vascular disease (PVD).
The compounds of this invention may be administered orally or parenterally, neat or in combination with conventional pharmaceutical carriers. Applicable solid carriers can include one or more substances which may also act as flavoring agents, lubricants, solubilizers, suspending agents, fillers, glidants, compression aids, binders or tablet-disintegrating agents or an encapsulating material. In powders, the carrier is a finely divided solid which is in admixture with the finely divided active ingredient. In tablets, the active ingredient is mixed with a carrier having the necessary compression properties in suitable proportions and compacted in the shape and size desired. The powders and tablets preferably contain up to 99% of the active ingredient. Any of the solid carriers known to those skilled in the art may be used with the compounds of this invention. Particularly suitable solid carriers include, for example, calcium phosphate, magnesium stearate, talc, sugars, lactose, dextrin, starch, gelatin, cellulose, methyl cellulose, sodium carboxymethyl cellulose, polyvinylpyrrolidone, low melting waxes and ion exchange resins.
Liquid carriers may be used in preparing solutions, suspensions, emulsions, syrups and elixirs of the compounds of this invention. The compounds of this invention can be dissolved or suspended in a pharmaceutically acceptable liquid carrier such as water, an organic solvent, a mixture of both or pharmaceutically acceptable oils or fat. The liquid carrier can contain other suitable pharmaceutical additives such as solubilizers, emulsifiers, buffers, preservatives, sweeteners, flavoring agents, suspending agents, thickening agents, colors, viscosity regulators, stabilizers or osmo-regulators. Suitable examples of liquid carriers for oral and parenteral administration include water (particularly containing additives as above e.g. cellulose derivatives, preferably sodium carboxymethyl cellulose solution), alcohols (including monohydric alcohols and polyhydric alcohols e.g. glycols) and their derivatives and oils (e.g. fractionated coconut oil and arachis oil). For parenteral administration, the carrier can also be an oily ester such as ethyl oleate and isopropyl myristate. Sterile liquid carriers are used in sterile liquid form compositions for parenteral administration.
Liquid pharmaceutical compositions which are sterile solutions or suspensions can be utilized by, for example, intramuscular, intraperitoneal or subcutaneous injection. Sterile solutions can also be administered intravenously. Compositions for oral administration may be either liquid or solid composition form.
Preferably, the pharmaceutical compositions containing the compounds of this invention are in unit dosage form, e.g. as tablets or capsules. In such form, the compositions may be sub-divided in unit doses containing appropriate quantities of the present compounds. The unit dosage forms can be packaged compositions, for example packeted powders, vials, ampoules, prefilled syringes or sachets containing liquids. Alternatively, the unit dosage form can be, for example, a capsule or tablet itself, or it can be the appropriate number of any such compositions in package form.
The therapeutically effective amount of the compounds of this invention that is administered and the dosage regimen depends on a variety of factors, including the weight, age, sex, and medical condition of the subject, the severity of the disease, the route and frequency of administration, and the specific compound employed, and thus may vary widely. However, it is believed that the pharmaceutical compositions may contain the compounds of this invention in the range of about 0.1 to about 2000 mg, preferably in the range of about 0.5 to about 500 mg and more preferably between about 1 and about 100 mg. Projected daily dosages of active compound are about 0.01 to about 100 mg/kg body weight. The daily dose can be conveniently administered two to four times per day.